Meat manufacturers are looking for ways to enable them to supply retail outlets from efficient, cost effective, central-processing centers. Increased shelf life with regard to both color (consumer acceptance) and spoilage (consumer safety) is required to make this possible as meat makes its way through longer distribution channels from producer to retailer to consumer.
Color shelf life is important to consumer acceptance. Consumers judge the freshness of meat by the presence of bright red oxymyoglobin pigment. Oxymyoglobin in fresh meat decreases with time during storage as it changes to the stable brown pigment, metmyoglobin. Although oxymyoglobin pigment fades during dark storage, for example in a meat locker, pigment loss is most pronounced in lighted, refrigerated display cases in retail establishments. Although pigment loss is primarily cosmetic in nature, it has serious economic consequences. Consumers in search of the freshest looking cuts avoid purchasing meat containing even small amounts of brown metmyoglobin. The unsaleable product which results from oxymyoglobin loss in red meats costs the industry an estimated $700 million dollars annually.
Shelf life associated with microbial spoilage is a more serious issue. The potential liability associated with food born illness outbreaks from the sale of microbially contaminated meat is enormous. The meat industry and associated retail outlets are seeking ways to insure consumer safety by preventing microbial contamination all along the manufacturing process. Process improvements such as carcass washing and carefully controlled low temperature processing are now routine in the industry. Modified atmosphere packaging (MAP) of meat products has also improved microbial shelf life of fresh meat products. Some processors have begun treating meat with ionizing radiation to extend the microbial shelf life of meat products. The irradiation process is an effective method for controlling microorganisms on meat, but many consumers are wary of its use. There is a need in the industry for antimicrobial methods and processes which are perceived by consumers as being more natural. The invention described below addresses this demand using GRAS (generally regarded as safe) seasonings.
Centrally processed meat will encounter at least two distinct storage environments prior to sale to a consumer. It will be stored in the dark at approximately 4° C. soon after production and during distribution. Prior to sale, it is likely to be stored in a refrigerated, illuminated display case. The general problem of enhancing the shelf life of fresh meat, then, can be separated into three subcategories: preserving the color during dark storage, preserving the color during storage in the lighted display case, and preventing the growth of spoilage organisms and pathogens throughout the commercially desirable storage period. We have discovered that treating meat with a combination of Labiatae herb extracts and hop extracts containing beta acids provides a novel method for enhancing the color shelf life under both dark and light storage conditions and for suppressing the growth of microorganisms for a commercially desirable period. The Labiatae herb extracts have been found to round out and suppress the bitter flavor of the hop extracts and allow for surprisingly high and truly effective concentrations of hop extracts to be added to meat without a negative flavor impact. The composition of this invention, a combination of hop extracts, Labiatae herb extracts and a storage atmosphere containing oxygen has been found to inhibit the growth of Gram positive microorganisms and very surprisingly, even Gram negative microorganisms. Prior art teaches that hop extracts do not control Gram-negative organisms. The hop and rosemary extracts are synergistic in their effects.
The antimicrobial activity of hop extracts and compounds against Gram-positive bacteria has been known for a long time. Hop extracts have not been considered effective against Gram-negative organisms. The antimicrobial activity of hop compounds has been studied mostly in growth media. Minimum inhibitory concentrations (MIC) have been determined in these media. For beta acids, the MIC is around 1 ppm. When tested in food, however, the MIC levels rise dramatically. The primary influences on MIC being fat content (the higher the fat content, the less antimicrobially active the hop acid). Another factor is pH (the lower the pH, the more active the hop acid). MICs for skim and 2% milk have been reported at 100 ppm. This rises to 1000 ppm in whole milk. The Millis et al patent teaches that beta acid flavor is noticeable at 15 ppm and becomes objectionable at 50 ppm. When hop acids are used at higher levels, the bitter flavor imparted to the food becomes a significant limiting problem. The main prior art is summarized below:
M. Teuber and A. F. Schmalreck (Arch. Mikrobiol. 94 (1973) pp. 159-171) review the use of hop extracts in medicine and reiterate that gram-negative microorganisms are generally not affected by hop extracts. Minimum inhibitory concentrations effective for Gram-positive organisms were determined for:
Lupulone (beta acids) 1 g/ml (1 ppm)humulone 2 g/ml (2 ppm)isohumulone 25 g/ml (25 ppm)humulinic acid250 g/ml (250 ppm)
W. J. Simpson and J. R. M. Hammond, (Antibacterial Action of Hop Resin Materials, EBC Congress, 1991, Chapter 21, pp. 185-192) describe the mode of action of trans-isohumulone (an isoalpha acid) and colupulone (a beta acid) against beer spoilage organisms. They indicate that low pH favors the antibacterial activity of isohumulone. The relative activity dropping from 226 at pH 3.8 to 42 at pH 4.6. They also demonstrate that colupulone has an effect on intracellular pH of recombinant lactic acid bacteria containing lux genes from the marine organism Vibrio fischeri. By implication, colupulone had antibacterial activity against this Gram-positive lactobacillus.
A. E. Larson, et al. (Antimicrobial Activity of Hop Extracts Against Listeria monocytogenes in Media and in Food, Int. J. Food Microbiol. 33 (1996) pp. 195-207) describe the effect of hop extracts containing varying amounts of humulones (alpha acids) and lupulones (beta acids) on controlling Listeria, a Gram-positive microorganism, in media and in certain foods. A hop extract (II) consisting of 41% beta acids, 12% alpha acids, and the remainder a mixture of desoxy-alpha acids, hop oils and hop waxes was found to be inhibitory at a concentration of 0.1 mg/ml (100 ppm) in skim and 2% milk and 1 mg/ml (1000 ppm) in whole milk and was listericidal in low fat cottage cheese at a concentration of between, 100 and 3000 ppm. A hop extract (III) consisting of 29.7% colupulone, 65% lupulone plus adlupulone, 8% desoxy-alpha acids, 7% water and 0.6% isoalpha acids enhanced the rate of inactivation of Listeria in coleslaw at a concentration of 1 mg/g (1000 ppm). Extract (III) showed no inhibitory effect even at 10,000 ppm in full fat camembert cheese. Both extracts (II) and (III) were inhibitory in trypticase soy broth cultures at the 0.01 mg/liter (0.01 ppm) level. This prior art teaches that inhibitory effects exhibited by hop extracts in media grossly over exaggerate the effectiveness of the hop extract in an actual food matrix. For example, the difference between 0.01 ppm in broth and 1000 ppm in coleslaw is a factor of 100,000. They conclude that because something works in culture media does not indicate it will work in food systems. Food systems require much higher concentrations of hop acids to show an antimicrobial effect than would have been predicted by the simple culture tests. This paper also teaches that “Overall, the antimicrobial activity of hop extracts appears to increase with acidity and lower fat content. Our results indicate that hop extracts could be used to control L. monocytogenes in minimally processed food with low fat content.”
E. A. Johnson and G. J. Haas (UK Patent Application GB 2,330,076, publication date Apr. 14, 1999) teaches that hop extracts are useful antibacterial agents against Clostridium botulinum and Clostridium difficile, both Gram positive organisms. They state that concentrations of 1 ppm or greater beta acids or hop extracts inhibit the growth of these organisms. Their examples are in the form of lab culture experiments in growth media. They surmise that hop extracts can be conveniently incorporated into food products to prevent disease caused by these microorganisms. This teaching is counter to the teaching by the same authors (A. E. Larson et al.) cited above which advises that extrapolations from culture media experiments to complex foods are not straightforward.
Millis et al. (U.S. Pat. No. 5,206,586) claims the use of beta acids for inhibiting Listeria in packaged foods, at a concentration of 6-50 ppm. Examples show culture only, not extension to food systems. Millis et al. claims there are serious flavor limitations to the use of beta and states that beta is noticeable at 15 ppm and objectionable above 50 ppm. Millis et al.'s claims on inhibition of Listeria in packaged foods are not supported by examples, but based upon his demonstrations of inhibition in broth. The Larson et al. prior art cited above shows that such a sweeping generalization is not supportable.
Barney et al., U.S. Pat. No. 5,455,038, teaches that tetrahydroisohumulone and hexahydrocolupulone are superior to Millis et al.'s beta acids for inhibiting Listeria in cultures. Tetrahydroisohumulone inhibited Listeria in soy broth at a concentration of between 6-18 ppm. Under the same conditions, hexahydrocolulupone inhibited Listeria at 0.4 ppm.
Barney et al., U.S. Pat. No. 5,370,863 teaches that tetrahydroisohumulone can inhibit Gram-positive bacteria that cause periodontal disease.
Maye, et al., PCT Application WO 00/52212 teaches that the acid form of the hop acid is superior to the salt form in inhibiting bacteria in an aqueous process stream. All hop acids appear to be covered.
Johnson and Haas, Japanese Patent Application 11-221064 teach the use of spraying foods or drinks with a solution (preferably an ethanol solution) with a hop extract or the ingredients of a hop extract in a concentration of >1 ppm and preferably at least 5 ppm, preferably further containing beta acid, preferably in the presence of a surfactant such as tween 80.
Rhodia Corporation has introduced a line of spray-dried solids containing hop ingredients added to control Gram positive organisms. They are extensions of a product line which use propionobacteria cultures as a natural source of propionates, which are well known antimicrobial compounds effective against Gram-negative organisms. Microgard® MG 225 consists of a dextrose based culture and a natural hop flavor. This product is especially effective against cold loving Gram-negative bacteria, certain yeasts and molds and select Gram-positive bacteria. Microgard® 325 consists of a skim milk based culture and natural hop flavor. It is reportedly active against Gram-positive organisms in low fat, low protein foods.
W. J. Simpson and A. R. W. Smith (J. Appl. Bacteriol. 72, (1992), pp. 327-334) showed that antibacterial activity increases with decreasing pH and that lipid material interferes with the activity of trans-isohumulone against Lactobacillus brevis, a Gram positive organism.
G. J. Haas and R. Barsoumian (J. Food Protect. 57, (1994) pp. 59-61) examined isoalpha acids and beta acids against a variety of microorganisms and looked at resistance development. Minimum inhibitory concentrations of isoalpha acids were in the 0.01 to 0.03% (100-300 ppm) in tryptic soy broth. MIC for beta acids were 0.003-0.01% (30-100 ppm) in the same media against a variety of Gram-positive Staphylococcus organisms. E. coli B, a Gram negative, was not sensitive to either of the hop resins.
J. S. Hough et al., (Brew. Ind. Res. Found. 63, (1957) pp. 331-333) provide another example of effectiveness against Gram positive organisms and ineffectiveness against Gram negative (Acetobacter suboxydans). MIC for lupulone=1-10 ppm in culture.
J. L. Shimwell (J. Inst. Brew. 43, (1937) 191-195) provides yet another example of activity against Gram positive and inactivity or even stimulating effects against Gram negative.
The antimicrobial activity of Labiatae herbs has also been the subject of study. Most prior art indicates that the antimicrobial activity of the herbs is centered in the volatile essential oil components.
P. M. Davidson and A. S. Naidu (in Natural Food Antimicrobial Systems, A. S. Naidu, ed., 2000, CRC Press, Boca Raton, pp. 265-294) review the antimicrobial properties of phyto-phenolic compounds from essential oils of spices, herbs, edible grains and seeds. The authors teach that the antimicrobial effects of spices and herbs are primarily due to the presence of phenolic compounds in the essential oil fractions and that some terpenes seem to show some activity, as well. Carvacrol, p-cymene and thymol are identified as the major volatile components of oregano, thyme and savory that likely account for the observed activity. The active antimicrobial agents of rosemary have been suggested to be borneol, camphor, 1,8-cineole, alpha pinene, camphene, verbenone and bornyl acetate. The active constituent of sage has been suggested to be thujone. Minimum lethal concentrations of essential oils of thyme oil have been shown to range from 225-900 ppm in cultures. These concentrations of essential oils in foods would cause serious flavor problems. Since culture experiments underestimate the concentration necessary for effectiveness in foods, the flavor problems in foods are likely to be more serious than even the culture numbers suggest. In another portion of this reference, minimum inhibitory concentrations of essential oils were stated as 1-2% for rosemary, 0.12-2% for thyme, 0.12-2% for spearmint, 0.5-2% for sage, 0.5-2% for peppermint and 0.12-2% for oregano. In the summary, the authors state that concentrations of antimicrobial compounds in herbs and spices are too low to be used effectively without adverse effects on the sensory characteristics of a food.
Y. Kimura et al., U.S. Pat. No. 4,380,506, teach a process for producing a preservative having antioxidant and antimicrobial activity. The process involves partitioning an extract of herb spices between polar and non-polar solvents. Some of the partitioned extracts showed antimicrobial activity against Gram positive Bacillus subtilis microorganisms in culture media. This reference does not anticipate the benefits of combining hop extracts with Labia tae herb as described in the present invention. The present invention does not require the partitioning process taught by Kimura et al and avoids the use of additional processing expense.
D. Ninkov (International Application WO 01/15680 A1) teaches that pharmaceutical compositions can be prepared by combining extracts of essential oils from plants of the Labiatae family with an organic acid or group 1 salt. Ninkov teaches that the antimicrobial activity of the pharmaceutical composition is due to the presence of organic phenols such as isopropyl o-cresol in the oil extract from the plant.
K. Shetty and R. G. Labbe (Asia Pacific J. Clin. Nutr. (1998, 7(3/4): 270-276.) describe work to clone Laminacae plants to produce enhanced levels of essential oil components such as carvacrol and thymol. These essential oil components have some antimicrobial properties but their commercial use is prevented by the strong flavors imparted to foods by these volatile compounds.
J. Campo, M. Amiot and C. Nguyen-the (2000, Journal of Food Protection 63, pp. 1359-1368) teach that rosemary extract has antimicrobial properties in culture studies. Minimum inhibitory concentrations varied with the species of bacteria being tested, but ranged from 0.06-1%. Using up to one percent of an ethanolic solution of rosemary had no effect on Gram negative bacteria. These researchers suggest that rosemary extract may show promise in foods with low fat and low protein content, but only against Gram positive organisms. No food systems were actually studied.
A. E. Down, et al., “Comparison of Vitamin E, Natural Antioxidants and Antioxidant Combinations on the Lean Color and Retail Case-Life of Ground Beef Patties” published in October, 1999, http://www.ansi.okstate.edu/research/1999rr/04.htm describes the effect of rosemary extract in combination with other natural antioxidants and vitamin E diet supplementation on the color life of non-MAP ground beef. The slight increase in color life observed using natural antioxidant blend containing rosemary is statistically indistinguishable from the control. This paper does not teach how to extend the microbial shelf life of meat.
A. E. Down, et al., “Influence of Vitamin E, Duralox®, and Herbalox® on Lean Color and Retail Case-Life of Ground Beef” published in October, 1999, http://www.ansi.okstate.edu/research/1999rr/05.htm, describes the effect of rosemary extract, rosemary extract in combination with other natural antioxidants and vitamin E on the color life of non-MAP ground beef. Addition of rosemary and rosemary plus other antioxidants increases the color life of the meat over the control, but is not as effective as addition of Vitamin E. This paper does not teach how to extend the microbial shelf life of meat.
Our studies in actual meat systems show that rosemary extract, Herbalox® Seasoning, in which the majority of the volatile oil components has been removed shows very little, if any, antimicrobial effect. Herbalox® is a registered trademark of Kalsec®, Inc.
None of the prior art on the antimicrobial use of rosemary or other Labiatae herbs either anticipates or renders obvious the present invention. The prior art focuses on the use of herb essential oils. The Labiatae herb extracts used in the present invention are processed in a manner that makes them essentially free of the native essential oil. The prior art neither anticipates nor renders obvious the synergistic combination of Labiatae herb extracts and hop extract. The prior art neither anticipates nor renders obvious the flavor masking effect of Labiatae herb extracts on the bitter flavor of the hop extracts. The prior art neither anticipates nor renders obvious the surprisingly beneficial antimicrobial effect of the combination of Labiatae herb extract, hop extract and high oxygen atmosphere packaging on both Gram positive and Gram negative organisms.